Kinetic analysis of nonphotochemical quenching of chlorophyll fluorescence. 1. Isolated chloroplasts

Nonphotochemical quenching of chlorophyll fluorescence in plants is indicat ive of a process that dissipates excess excitation energy from the light-ha rvesting antenna of photosystem II. The major fraction of quenching is obli gatorily dependent upon the thylakoid Delta pH and is regulated by the deep oxidation state of the xanthophyll cycle carotenoids associated with the li ght-harvesting complexes. Basic principles of enzyme kinetics have been use d to investigate this process in isolated chloroplasts. The extent of quenc hing was titrated against the estimated thylakoid lumen pH, and a sigmoidal relationship was obtained with a Hill coefficient of 4.5 and a pK of 4.7. Upon de-epoxidation, these parameters changed to 1.6 and 5.7, respectively. Antimycin A suppressed quenching, increasing the Hill coefficient and redu cing the pK. The rate of induction of quenching fitted second-order kinetic s with respect to illumination time, and the rate constant was dependent up on the Delta pH, the de-epoxidation state, the presence of antimycin, and a lso the presence of dibucaine, a quenching enhancer. All these data are con sistent with the notion that quenching is caused by a conformational transi tion in a chloroplast thylakoid protein; this transition shows cooperativit y with respect to proton binding, and is controlled by de-epoxidation state and various exogenous reagents.